Structural features and stability of an RNA triple helix in solution

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• RNA can form complex secondary and tertiary structures beyond simple base pairing.
• Intramolecular triple helices represent a less explored structural motif in RNA.

Purpose of the Study:

• To characterize the structure and stability of a de novo designed RNA intramolecular triple helix.
• To elucidate the nature of base pairing and stabilizing interactions within the RNA triple helix.
• To provide insights into RNA structural diversity and its implications for molecular design.

Main Methods:

• One- and two-dimensional Nuclear Magnetic Resonance (NMR) spectroscopy to determine atomic-level structure.
• UV absorption measurements to assess thermal stability and melting transitions.
• Computational model building guided by NMR data.

Main Results:

• The 30-nucleotide RNA sequence formed a stable intramolecular triple helix with seven pyrimidine-purine-pyrimidine base triples.
• Watson-Crick and Hoogsteen base pairing interactions were identified as key stabilizing forces.
• The triple helix exhibited high stability in acidic pH (4.3), melting at 62°C in a single transition.
• NMR data supported a structural model involving an A-helix formation by Watson-Crick paired strands.
• A potential hydrogen bond was proposed between the 2' hydroxyl of the Hoogsteen strand and a phosphate oxygen.

Conclusions:

• The designed RNA triple helix is a stable structure stabilized by combined Watson-Crick and Hoogsteen interactions.
• The stability is pH-dependent, with enhanced stability observed at acidic pH.
• The findings contribute to understanding RNA structural plasticity and inform the design of RNA-targeting molecules.